Characterization of the procoagulant phenotype of AT-deficient mice. (A) Schematic diagram of experimental design, in which 6- to 8-week-old C57BL/6 mice were treated with IV injection of AT siRNA (3.5 μg/g bodyweight), complexed with Invivofectamine 3.0. After 24 and 48 hours of siRNA injection, AT variants (AT-WT, AT-4Mut, and AT-R425del) were administered IV at 500 μg per injection. Mice were euthanized 72 hours after siRNA injection. (B) Expression of AT gene in liver tissues were measured through quantitative real-time polymerase chain reaction after 48 and 72 hours of siRNA treatment. β-actin was used as the reference gene. (C) Plasma levels of AT in control siRNA and AT siRNA–treated mice were measured by ELISA after 48 and 72 hours of the siRNA treatment. (D) Plasma levels of AT at the 72-hour time point were measured by ELISA. (E) Frequency of periocular hemorrhage for different animal groups in female mice is presented. (F) Morbidity scores in different animal groups are determined as described in “Methods.” (G) aPTT in different animal groups was measured using Stago ST4 coagulation analyzer. (H) Plasma clotting time in different animal groups was measured using STart 4 coagulation analyzer. (I) Plasma fibrinogen level in different animal groups was measured by ELISA. (J) Peripheral blood counts were measured using Hemavet 950 veterinary hematology analyzer. Platelet counts of different animal groups are presented. All data are presented as mean ± standard error of the mean (SEM). n ≥ 7. Statistical analysis was performed using Graph Pad Prism 10. P values are determined by 1-way analysis of variance (ANOVA), followed by Bonferroni multiple comparison test. ∗P < .05; ∗∗P < .01; ∗∗∗P ≤ .001; ∗∗∗∗P ≤ .0001. ns, not significant.